Comparative performance analysis of different microfilaria testing methods for Dirofilaria immitis in canine blood.

Background: Microfilaria (MF) testing is an essential part of canine heartworm diagnostics, and it is recommended by the American Heartworm Society that a MF test be performed in tandem with antigen testing on every dog, every year, regardless of prevention status or history. There are a variety of methods that can be used to detect MF in canine whole blood; however, these methods widely vary in their sensitivities as well as practical factors, including time investment and cost. Additionally, some MF tests offer the advantage of being quantitative or allowing for morphological or molecular species identification, while other tests should only be used qualitatively.

Methods: The purpose of this study is to evaluate the quantitative and qualitative performance of MF tests, including the 20 μL count, wet mount, 9 μL and 40 μL hematocrit tubes, thin smear, thick smear, modified Knott test (MKT), and conventional polymerase chain reaction (PCR).

Results: Qualitatively, there was little difference in the performance of the 20 μL count, wet mount, MKT, and PCR. The MKT and PCR are the optimal MF tests, as these perform most reliably for detecting positives even when the MF per milliliter is relatively low, and in most cases, these two methods also allow for species-level confirmation of the identity. However, PCR tends to be a very costly test, and both PCR and MKT require a greater degree of expertise and time investment to perform than other tests. Even the lowest performance tests, including the thin smear and hematocrit tube methods, can reliably detect MF at very high burdens; although, caution should be advised when using low reliability methods, since there is a greater likelihood of failing to identify MF-positive dogs.

Conclusions: Microfilaria (MF) testing is an essential part of heartworm diagnosis and screening in dogs, and test selection should balance practical factors such as cost and time investment with the patient's risk of infection based on prevention status and history, clinical signs, and antigen testing results. This approach to MF testing will help minimize cost while avoiding failure to detect MF in infected dogs, especially when MF burden is low.